The goal of this exploratory project (R21) is to identify therapeutic vaccine candidates that can be used to prevent tuberculosis (TB) in hosts latently infected with Mycobacterium tuberculosis. We have developed a mouse model of TB in which latent infection can be reproducibly induced to progress to disease without immunologically manipulating the animal. We previously showed that a strain of M. tuberculosis disrupted in a 13-gene operon called mcel is unable to elicit a pro-inflammatory response in mice that would otherwise allow the organism to enter a latent state of infection in the lungs. Disruption of the operon's putative negative regulator called mcelR leads to dysregulation of the immune response in mice infected with the mutant strain. The mouse infected with the mcelR mutant mounts an uncontrolled inflammatory response--opposite of the response observed with the mcel operon infection. In the Cornell model of infection, the mcelR mutant can reproducibly cause disease in mice within 1 month after cessation of antibiotics, without the administration of steroids. We wish to, therefore, use this mutant to identify M. tuberculosis products that will prevent disease production following cessation of antibiotics. The aims of the project are to 1) optimize the mouse Cornell model to develop a protocol that can be used for a therapeutic vaccine trial in mice, and 2) identify M. tuberculosis products that prevent TB using the selected protocol for the vaccine trial. We will identify the optimal inoculum dose administered by an aerosol route that will reproducibly produce disease in latently infected BALB/c mice. We will also identify the optimal time period in which disease manifests after a period of latency. Once these conditions are established, this protocol will be used to screen M. tuberculosis products that will prevent this disease manifestation caused by the mcelR mutant. Such products will then serve as therapeutic vaccine candidates. If we do identify such products, we will seek additional funding for a long-term project to evaluate such products in nonhuman primates in collaboration with those involved in such animal work. Therapeutic vaccines against TB is a major priority in TB control, and we have an opportunity to identify such products by a novel approach using a simple, inexpensive, and reproducible animal model.